Project Summary Malaria remains a significant contributor to the global burden of disease. In endemic regions, malaria mostly affects children. Older individuals are protected from disease by an immune response against the parasite?s blood stage. Eliciting a protective immune response in children by vaccination would significantly reduce malaria morbidity and mortality. Antibodies are a critical component of such protection. A strong correlation exists between antibodies against merozoite antigens and a reduction in malaria incidence. Unfortunately, vaccines based on these antigens have thus far not been effective, suggesting that our understanding of protective immunity is far from complete. This proposal stems from the observation that persistent malaria exposure eventually gives rise to antibodies reactive against heterologous parasite strains. The long period of time necessary for the development of protective anti-malaria immunity is suggestive of a long evolutionary pathway towards the generation of such cross-strain reactive antibodies. In addition, protection against disease has been shown to correlate with selected antibody effector functions. We hypothesize that the memory B cell pool of malaria-exposed individuals will contain cells that make cross-strain reactive IgGs that are directed against merozoite antigens, are highly somatically mutated and are of selected IgG subclasses. To test this hypothesis, we will compare structural and functional features of antibodies directed against two merozoite antigens, MSP1 and AMA1, among immune adults, semi-immune children, and children who have been exposed but are still susceptible to malaria. In Specific Aim 1, we will isolate MSP1- and AMA1-specific memory B cells from these individuals and express their monoclonal antibodies using linear expression cassettes to confirm antigen specificity and determine cross-strain reactivity. In Specific Aim 2, the heavy and light chain variable region sequences and IgG subclasses will be determined for a comparative analysis of antibody structures among the three groups of individuals. In addition, antibodies with diverse structural features will be cloned and expressed in their original IgG subclass, and their inhibitory capacity against four different strains of P. falciparum will be analyzed in functional assays. This project will use valuable material from naturally exposed individuals in combination with advanced methodologies to dissect protective anti-malaria immunity. The comparison of structural and functional antibody features between susceptible and protected individuals will provide a wealth of data about the types of antibodies that a blood stage vaccine should ideally elicit, with respect to features of both the ?head? and the ?tail? of these antibodies. In a follow-up study, identification of the conserved epitopes recognized by these antibodies will reveal novel targets for immunization.